Marintchev, A. | bioRxiv Science

bioRxiv2024

Molecular basis for the interactions of eIF2β with eIF5, eIF2B, and 5MP1 and their regulation by CK2

The heterotrimeric GTPase eukaryotic translation initiation factor 2 (eIF2) delivers the initiator Met-tRNAi to the ribosomal translation preinitiation complex (PIC). eIF2{beta} has three lysine-rich repeats (K-boxes), important for binding to the GTPase-activating protein eIF5, the guanine nucleotide exchange factor eIF2B, and the regulator eIF5-mimic protein (5MP). Here, we combine X-ray crystallography with NMR to understand the molecular basis and dynamics of these interactions. The crystal structure of yeast eIF5-CTD in complex with eIF2{beta} K-box 3 reveals an extended binding site on eIF2{beta}, far beyond the K-box. We show that eIF2{beta} contains three distinct binding sites, centered on each of the K-boxes, and human eIF5, eIF2B{varepsilon}, and 5MP1 can bind to all three sites, while reducing each others affinities. Our results reveal how eIF2B speeds up the dissociation of eIF5 from eIF2-GDP to promote nucleotide exchange; and how 5MP1 can destabilize eIF5 binding to eIF2 and the PIC, to promote stringent start codon selection. All these affinities are increased by CK2 phosphomimetic mutations, highlighting the role of CK2 in both remodeling and stabilizing the translation apparatus.

biochemistry↗

bioRxiv2022

The PCI domains are "winged" HEAT domains

The HEAT domains are a family of helical hairpin repeat domains, composed of four or more hairpins. HEAT is derived from the names of four family members: huntingtin, eukaryotic translation elongation factor 3 (eEF3), protein phosphatase 2 regulatory A subunit (PP2A), and mechanistic target of rapamycin (mTOR). HEAT domain-containing proteins play roles in a wide range of cellular processes, such as protein synthesis, nuclear transport and metabolism, and cell signaling. The PCI domains are a related group of helical hairpin domains, with a "winged-helix" (WH) subdomain at their C-terminus, which is responsible for multi-subunit complex formation with other PCI domains. The name is derived from the complexes, where these domains are found: the Proteasome "lid" regulatory subcomplex, the COP9 signalosome (CSN), and eukaryotic translation initiation factor 3 (eIF3). We noted that in structure homology searches using HEAT domains, sometimes PCI domains appeared in the search results ahead of other HEAT domains, which indicated that the PCI domains could be members of the HEAT domain family, and not a related but separate group. Here, we report extensive structure homology analysis of HEAT and PCI domains, both within and between the two groups of proteins. We present evidence that the PCI domains as a group have greater structural homology with individual groups of HEAT domains than some of the HEAT domain groups have among each other. Therefore, our results indicate that the PCI domains have evolved from a HEAT domain that acquired a WH subdomain. The WH subdomain in turn mediated selfassociation into a multi-subunit complex, which eventually evolved into the common ancestor of the Proteasome lid/CSN/eIF3.

bioinformatics↗